Gustatory preference under redlight

Gustatory preference under bluelight

It’s not yet confirmed, that the 3IY treatment is working

To see whether the initial preference but also the preference shift, observed for flies expressing the optogenetic Chrimson-Channel in neurons from the PPM2 cluster (tested for preference in JoyStick and T-Maze), does depend on dopamine signaling, I want to repeat these experiments with dopamine depleted flies. For this I plan to use 3-Iodo-Tyrosine, which should reduce dopamine production. To verify the effect 3IY I conducted two sets of experiments, testing locomotion and geotaxis. Successfully depleting dopamine should decrease locomotion and performance in geotaxis assays. At this point both experiments have the same outcome, that the tested method to apply 3IY does not work.

Locomotion in open field arena

Geotaxis in Benzer Counter-Current

Gustatory preference under redlight

Gustatory preference under redlight

CantonS under bluelight

Preference Index: (Pur-Salz)/(Pur+Salz+Neutral)

Stichprobengröße N: 20 jeweils

CantonS under redlight

Preference Index: (Pur-Salz)/(Pur+Salz+Neutral)

Stichprobengröße N: 20 jeweils

Bluelight

PI: (Pur-Salz)/(Pur+Salz+Neutral)

Stichprobengröße: N(0M) = 12; N(0,025M) = 4; N(0,5) = 12; N(1,5M) = 13; N(2,5M) = 12; N(3,5M) = 10

Redlight

PI: (Pur-Salz)/(Pur+Salz+Neutral)

Stichprobengröße: N(0M) = 14; N(0,025M) = 4; N(0,5) = 13; N(1,5M) = 12; N(2,5M) = 13; N(3,5M) = 10

Stichprobengröße pro Gruppe: N = 10

PI: (Pur-Salz)/(Pur+Salz+Neutral)

Stichprobengröße pro Gruppe: N = 10 und bei 0M N = 15

PI:(Pur-Salz)/(Pur+Salz+Neutral)

To investigate whether there are subsets of dopaminergic neurons that are responsible for “reward” or “punishment” in the absence of stimuli, different transgenic Drosophila lines were screened for approach or avoidance of neuronal activation. The different lines expressed the optogenetic “Chrimson” channel in different sets of dopaminergic neurons and were tested in different operant self-stimulating experiments (T-Maze (red light), T-Maze (yellow light), Y-Maze and JoyStick). Except for two, none of the tested lines displayed consistent preference or avoidance in all four different experiments. Both lines (TH-D’ and TH-D1) drive expression in the same DAN clusters including PPL1_FB, PPM2 and PPM3.

Since there are now more refined Gal4 driver lines, enabling expression of the optogenetic channel in only subsets of neurons compared to the original driver lines, T-Maze and JoyStick experiments were repeated with these new lines.

Rescreening in JoyStick and T-Maze showed that out of the three tested lines only one line displayed (tendencies for) light-avoidance as the original driver line. This was the line driving expression in the PPM2 cluster. (JoyStick data from other lines not shown, PPM3 could not be tested in T-Maze due to technical limitations). Interestingly in the JoyStick, this line displayed a shift in preference that was similar to the shift observed in the original TH-D’ driver line.

As mentioned above, JoyStick experiments had shown that after initial avoidance of the light (represented by negative PIs), in the last training periods flies would show PIs close to zero. In the T-Maze experiments flies are usually tested for only 1 minute. To find out whether fly preference would show a same development when choice time was extended, I conducted a set of T-Maze experiments, with a choice time of 10 minutes.

These results suggest that in the flies expressing the Chrimson channel in PPM2 DANs, prolonged choice time leads to a shift in “valence”, as also observed in the broadly expressing driver line TH-D’. This shift was only observed under yellow light.

Comparing control groups in yellow and red light, there seem to be differences in the activation of the Chrimson channel under red and yellow light. Whether this can be explained by different penetration capabilities due to the different wave lengths will be subject to further research.